1. Field of the Invention
The present invention relates to methods for preparing hydrophobic drug loaded hydrophilic microspheres, having a non-burst and sustained release local delivery of drug at the site of embolisation.
2. Description of the Related Art
Embolisation therapy involves the introduction of an agent into the vasculature in order to bring about the deliberate blockage of a particular vessel. This type of therapy is particularly useful for blocking abnormal connections between arteries and veins (such as arteriovenous malformations, or AVMs), and also for occluding vessels that feed certain hyper-vascularised tumours, in order to starve the abnormal tissue and bring about tumour ischemia or necrosis.
The process of embolisation may induce tumour necrosis or ischemia depending upon the extent of the embolisation. The response of the tumour cells to the hypoxic environment can result in an ensuing angiogenesis in which new blood vessels are grown to compensate for the loss of flow to the tumour by the embolisation. It would be desirable therefore to combine embolisation with the administration of agents that could prevent the ensuing angiogenic response or combine with the release of a cytotoxic or other anti-tumoral agent to bring about cell death in those cells that are not killed by the embolisation.
In the early 1960s, the National Cancer Institute (NCI) in the United States initiated a programme of biological screening of extracts taken from a wide variety of natural sources. One of these extracts was found to exhibit marked antitumour activity against a broad range of rodent tumours. Although this discovery was made in 1962, it was not until five years later that two researchers, Wall and Wani, of the Research Triangle Institute, North Carolina, isolated the active compound, from the bark of the Pacific yew tree (Taxus brevifolia). In 1971, Wall and Wani published the structure of this promising new anti-cancer lead compound, a complex poly-oxygenated, Wani, M. C., H. L. Taylor, Monroe Wall, P. Coggon, A. T. McPhail, 1971, “Plant Antitumor Agents. VI. The Isolation and Structure of Taxol, a Novel Antileukemic and Antitumor Agent from Taxus brevifolia,” Journal of the American Chemical Society, 93: 2325-2327.
Paclitaxel is a natural product with antitumor activity. It is used to treat ovarian cancer, Karposi's sarcoma, and used in combinations with other chemotherapy agents to treat breast cancer, non-small cell lung cancer and is most effective against ovarian carcinomas and advanced breast carcinomas. Paclitaxel is given intravenously (it irritates skin and mucous membranes on contact). Paclitaxel, which is sold as Taxol® by Bristol-Myers Squibb, is obtained via a semi-synthetic process from Taxus baccata. The chemical name is 5β,20-Epoxy-1,2α,4,7β,10β,13α-hexahydroxytax-11-en-9-one 4,10-diacetate-2-benzoate 13-ester with (2R,3S)—N-benzoyl-3-phenylisoserine. Paclitaxel is a white to off-white crystalline powder with the empirical formula CH47H51NO14 and a molecular weight of 853.9. Paclitaxel is highly lipophilic, insoluble in water, and melts at around 216-217° C.
The relatively non-toxic properties of paclitaxel have made it a leading light in the treatment of cancer in the 1990s, providing a non-intrusive alternative to the more radical techniques of radiotherapy and surgery.
Despite its well-documented biological activity, very little interest was shown in paclitaxel until scientists at the Albert Einstein Medical College reported that its mode of action was totally unique. Until this finding in 1980, it was believed that the cytotoxic properties of paclitaxel were due to its ability to destabilise microtubules, important structures involved in cell division (mitosis). In fact, paclitaxel was found to induce the assembly of tubulin into microtubules, and more importantly, that the drug actually stabilises them to the extent that mitosis is disrupted. Such a novel mode of action was believed to make paclitaxel a prototype for a new class of anticancer drugs. Paclitaxel binds to microtubules and inhibits their depolymerization (molecular disassembly) into tubulin. It blocks a cell's ability to break down the mitotic spindle during mitosis (cell division). With the spindle still in place the cell cannot divide into daughter cells (this is in contrast to drugs like colchicine and the Vinca alkaloids, which block mitosis by keeping the spindle from being formed in the first place).
Most of the reported work on the preparation of paclitaxel-loaded polymeric drug delivery systems is based on hydrophobic polymer systems in which paclitaxel has good solubility.
WO 2003/077967 relates to a deposition method for applying an active substance to an endoprosthesis having a thin polymer coating. The deposition method enables a slow and largely constant administering of an active substance, as cited in the example of tretinoin. Since additional processing steps are not required after the application of the active substance(s), it is unnecessary to worry about coating conditions causing the active substance to be broken down, for example, by the application of second polymer coating. Even relatively unstable active substances, e.g. tretinoin, can be applied without any difficulties to the endoprosthesis. Thus 4-amino-[2,2]-paracyclophane was cleaved at 700° C., 20 Pa to reactive monomers and polymerised at the surface of a stent at 20° C. The polymer-coated stent was contacted with a DMSO solution of tretinoin and dipped into water; this resulted the precipitation of tretinoin onto the surface of the stent and embedding of the precipitate into the polymer layer.
Angiotech's group have studied the paclitaxel loading into poly(L-lactic acid) (PLLA) microspheres using solvent evaporation method. PLLA and paclitaxel were dissolved in dichloromethane. The organic phase was added to an aqueous solution of 2.5% poly (vinyl alcohol) under stirring. Subsequently, after 2 hr the aqueous suspension containing microspheres was passed through sieves to retain the particles in certain size ranges. The microspheres were further dried for 12-16 hr at ambient temperature. [Richard T. Liggins, Helen M. Burt Paclitaxel loaded poly(L-lactic acid microspheres: properties of microspheres made with low molecular weight polymers' International Journal of Pharmaceutics 222 (2001)19-33; Richard T. Liggins, Helen M. Burt ‘Paclitaxel loaded poly(L-lactic acid) microspheres II. The effect of processing parameters on microsphere morphology and drug release kinetics’ International Journal of Pharmaceutics 281 (2004) 103-106.] Later they extended their work on poly(lactic-co-glycolic acid) films for delivery of paclitaxel. [John K. Jackson, et al. ‘Characterization of perivascular poly(lactic-co-glycolic acid] films containing paclitaxel’ International Journal of Pharmaceutics 283 (2004)97-109.] Other work includes PEG-coated poly(lactic acid) microspheres. [Gladwin S. Das, et al. ‘Controlled delivery of taxol from poly(ethylene glycol)-coated poly(lactic acid) microspheres’ Journal of Biomedical Materials Research 55 (2001)96-103].
Boston Scientific Corporation has developed the system of coronary stent coating for delivery of paclitaxel by formulating polymer blends with 10 to 25% of paclitaxel. The polymers used are poly(butyl methacrylate), poly(styrene-co-isobutylene-co-styrene), or poly(styrene-co-(ethylene-butylene)-co-styrene), which are blended with poly(styrene-co-maleic anhydride). A recent development uses a modified styrenic portion, i.e. hydroxystyrene or its acetylated version. [Shrirang Ranade, et al. Abstracts of Papers, 229th ACS National Meeting, San Diego, Calif., US, Mar. 13-17, 2005, PMSE-022].
Composition and methods for in vivo controlled release of pharmaceutically active agents associated with hydroxyapatite (HAP) in a pharmaceutically acceptable carrier are described in WO 2003/030943. The pharmaceutically acceptable carrier can be a polymer paste or gel which may contain a second pharmacologically active agent. Methods of making and administering controlled release compositions for the delivery of a pharmacologically active agent, such as a nucleic acid, in combination with a polycationic polymer and in a pharmaceutically acceptable carrier, to a mammal in a pharmaceutically effective amount are provided.
Rapamycin, also known as sirolimus, was isolated the first time in 1969 from a fungus (Streptomyces hygroscopicus) in the island of Rapa Nui (Easter Island). Initially it was found to have potent antifungal and antiproliferative activities; but it was in 1977 when Martel et al reported its promising immunosuppressive activity [Martel, R. R.; Canadian Journal of Physiological Pharmacology, 55, 48-51 (1977).] From this time its mechanism of action has been thoroughly studied, and it is known how this antibiotic exerts its immunosuppressive and antiproliferative activities. Rapamycin is a white to off-white powder and is insoluble in water, but freely soluble in benzyl alcohol, chloroform, acetone, and acetonitrile.
Rapamycin and rapamycin analogues are currently in clinical development against a number of cancer indications. The mechanism of action is as an inhibitor of the mammalian target of rapamycin (mTOR). The cyclic macrolide structure inhibits cellular proliferation by interfering with the highly conserved TOR pathway, which control the synthesis of essential proteins involved in cell cycle progression.
mTOR is a protein kinase with similarities to the catalytic domains of phosphoinositide 3-kinases (PI3-k). Once activated, TOR transduces signals that initiate synthesis of ribosomal proteins, translation of specific mRNAs and generation of cyclin-dependent kinases, promoting the progression of the cell cycle. This results in activation and proliferation of T and B-cells and antibody production as well as proliferation of non-immune cells such as hepatocytes, fibroblasts, endothelial cells and smooth muscle cells. [Neuhaus P, Klupp J, Langrehr J M.; Liver Transpl. 2001 June; 7(6):473-84. mTOR inhibitors: an overview.]
Rapamycin exerts its antiproliferative effect mainly by blocking all of these events, as a consequence of inhibition of mTOR. It is able to inhibit this protein kinase by forming a trimeric stable complex, after binding with the soluble intracellular receptor protein FKBP12. This inhibition blocks the synthesis of cyclin-dependent kinases, which are key mRNAs that code for proteins required for cell cycle progression from G1 to S phase.
mTOR is also a positive regulator of hypoxia-inducible factor-1-dependent gene transcription in cells exposed to hypoxia or hypoxia mimetic agents [Hudson C C, Liu M, Chiang G G, Otterness D M, Loomis D C, Kaper F, Giaccia A J, Abraham R T.; Mol Cell Biol. 2002 October; 22(20):7004-14. Regulation of hypoxia-inducible factor 1alpha expression and function by the mammalian target of rapamycin.] If rapamycins prove to be effective inhibitors of hypoxic adaptation in developing tumours, these drugs could have dramatic effects on tumour growth, invasiveness and metastatic potential in cancer patients. In embolisation a hypoxic environment is induced and therefore rapamycin and its analogues may act mechanistically by inhibiting mTOR and consequently inhibiting the production of hypoxia induced factor (HIF-1) widely believed to be involved in angiogenic responses.
Treatment of tumour-bearing animals with rapamycin results in decreased expression of VEGF mRNA and decreased circulating levels of VEGF protein. Thus, proliferation of smooth muscle and endothelial cells is inhibited by mTOR inhibition. This anti-angiogenic effect may contribute to the efficacy of mTOR inhibitors in cancer therapy [Rao R D, Buckner J C, Sarkaria J N.; Curr Cancer Drug Targets. 2004 December; 4(8):621-35. Mammalian target of rapamycin (mTOR) inhibitors as anti-cancer agents].
Rapamycin and rapamycin analogues have demonstrated activity against a broad range of human cancers growing in tissue culture and in human tumor xenograft models. The central role of mTOR in modulating cell proliferation in both tumour and normal cells and the importance of mTOR signalling for the hypoxic response suggest that rapamycin-based therapies may exert anti-tumour effects primarily through either inhibition of tumour cell proliferation or suppression of angiogenesis. Although rapamycin can induce apoptosis in select tumour models, rapamycin treatment typically slows growth but does not induce tumour regression, suggesting that tumour cell loss through apoptosis or other mechanisms are not major contributors to drug effect in most cases.
There have been many reports of drug delivery systems using hydrophobic polymers, such as poly(L-lactic acid), poly(lactic-co-glycolic acid), poly(caprolactone), polybutyl methacrylate, and poly(styrene-co-isobutylene-co-styrene). However, there are few reports of hydrogel microspheres loaded with paclitaxel. This is due to the poor compatibility between hydrophobic drugs and hydrogel microspheres [R. Shi, H. M. Burt, ‘Amphiphilic dextran-graft-poly(epsilon-caprolactone) films for the controlled release of paclitaxel’ International Journal of Pharmaceutics 271 (2004) 167, http://www.ptca.org/articles/taxus_profileframe.html D. S. Das, G. H. R. Rao, R. F. Wilson, T. Chandy, ‘Controlled delivery of taxol from poly(ethylene glycol)-coated poly(lactic acid) microspheres’ Journal of Biomedical Materials Research, 55 (2001) 96 R. T. Liggins, H. M. Burt, ‘Paclitaxel loaded poly(L-lactic acid) microspheres: properties of microspheres made with low molecular weight polymers’ International Journal of Pharmaceutics, 222 (2001) 19. J. K. Jackson, J. Smith, K. Letchford, K. A. Babiuk, L. Machan, P. Signore, W. L. Hunter, K. Wang, H. M. Burt, ‘Characterisation of perivascular poly(lactic-co-glycolic acid) films containing paclitaxel’ International Journal of Pharmaceutics, 283 (2004) 97. S. K. Dordunoo, J. K. Jackson, L. A. Arsenault, A. M. C. Oktaba, W. L. Hunter, H. M. Burt, ‘Taxol encapsulation in poly(epsilon-caprolactone) microspheres’ Cancer Chemother. Pharmacol. 36 (1995) 279.]
US 2003/202936 discloses a process in which microspheres are prepared by immersing microparticles in a solution containing methanol and aminoacridine. Excess methanol is removed by evaporation, but this results in precipitation of the aminoacridine both inside and outside the microspheres.
Vandelli et al in the Journal of Controlled Release, 96 (2004), 67-84 disclose microspheres in which diclofenac is precipitated in the core. The drug is uniformly distributed in each microparticle. The presence of drug on or close to the surface leads to rapid initial release of the drug, which is often undesirable.